1. Field of the Invention
The invention relates generally to the fields of immunology and inflammatory bowel disease and more specifically to the diagnosis and treatment of a clinical subtype of ulcerative colitis.
2. Background Information
Inflammatory bowel disease (IBD) is the collective term used to describe two gastrointestinal disorders of unknown etiology: Crohn""s disease (CD) and ulcerative colitis (UC). The course and prognosis of ulcerative colitis, which occurs world-wide and is reported to afflict as many as two million people, varies widely. Onset of ulcerative colitis is predominantly in young adulthood with diarrhea, abdominal pain, and fever the three most common presenting symptoms. The diarrhea may range from mild to severe and often is accompanied by bleeding. Anemia and weight loss are additional common signs of UC. Ten percent to fifteen percent of all patients with inflammatory bowel diseases such as UC will require surgery over a ten year period. In addition, patients with UC are at increased risk for the development of intestinal cancer. Reports of an increasing occurrence of psychological problems, including anxiety and depression, are perhaps not surprising symptoms of what is often a debilitating disease that strikes people in the prime of life.
Unfortunately, the available therapies for ulcerative colitis are few, and both diagnosis and treatment have been hampered by a lack of knowledge regarding the etiology of the disease. What is clear, however, is that the pathogenesis of ulcerative colitis involves immune-mediated damage to the intestinal mucosa. Autoantibodies, specifically antibodies against cytoplasmic components of neutrophils (pANCA), have been reported in 68-80% of patients with ulcerative colitis, further supporting a role for immune dysregulation in this disease. However, the antigens recognized by these pANCA autoantibodies, which would be useful in diagnosing and treating UC patients have, to date, escaped identification.
In other inflammatory bowel diseases such as Crohn""s disease, bacteria have been implicated in the initiation or progression of the disease. That microbes can play a role in Crohn""s disease is supported, for example, by the efficacy of antibiotics and diet in mitigating disease in some Crohn""s patients. However, until now a role for microbes or microbial antigens in the immune dysregulation producing UC has not been suspected. Such microbial antigens can be the original inducers of the disease-related immune response in UC and, as such, can contain displayed B-cell epitopes that react especially effectively with UC pANCA autoantibodies. As a consequence, these microbial antigens can increase the level of UC serodetection from its current level of 60-70% with the fixed neutrophil assay. Such microbial antigens also can bear a disease related T-cell epitope and, as likely original inducers of the disease-related immune response, can be particularly effective tolerogenic antigens for treating UC patients.
Thus, there is a need for identification and isolation of UC pANCA target antigens, including antigens of microbial origin such as those expressed in colonic bacteria of UC patients. Such antigens would be useful for diagnosing and treating the large population of UC patients that have pANCA autoantibodies. The present invention satisfies this need by providing the histone H1 UC pANCA target antigen and additional UC pANCA microbial antigens. Related advantages are provided as well.
The present invention provides methods of diagnosing ulcerative colitis (UC) by obtaining a sample from a subject suspected of having inflammatory bowel disease; contacting the sample with a histone H1-like antigen, or pANCA-reactive fragment thereof, under conditions suitable to form a complex of histone H1-like antigen, or pANCA-reactive fragment thereof, and antibody to histone H1-like antigen; and detecting the presence or absence of the complex, where the presence of the complex indicates that the subject has ulcerative colitis. A histone H1-like antigen useful in these methods can be, for example, a protein immunoreactive with NANUC-2 and having an amino acid sequence having at least 65% amino acid identity with SEQ ID NO: 27.
The invention also provides methods of inducing tolerance in a pANCA-positive patient with UC by administering to the patient an effective dose of a histone-like H1 antigen, or tolerogenic fragment thereof. Such a histone H1-like antigen can be, for example, a protein immunoreactive with NANUC-2 and having an amino acid sequence having at least 65% amino acid identity with SEQ ID NO: 27.
The present invention also provides a composition of histone H1-like antigen, or tolerogenic fragment thereof, combined with a tolerogizing molecule. In such a composition, a histone H1-like antigen can be, for example, a protein immunoreactive with NANUC-2 and having an amino acid sequence having at least 65% amino acid identity with SEQ ID NO: 27.
In addition, the present invention provides methods of UC by obtaining a sample from a subject suspected of having inflammatory bowel disease; contacting the sample with a porin antigen, or pANCA-reactive fragment thereof, under conditions suitable to form a complex of porin antigen, or pANCA-reactive fragment thereof, and antibody to porin antigen; and detecting the presence or absence of the complex, where the presence of the complex indicates that the subject has ulcerative colitis. A porin antigen useful in these methods can be, for example, a protein immunoreactive with NANUC-2 and having at least 65% amino acid identity with SEQ ID NO: 28, SEQ ID NO: 29 or SEQ ID NO: 30.
The invention also provides methods of inducing tolerance in a pANCA-positive patient with UC by administering to the patient an effective dose of a porin antigen, or tolerogenic fragment thereof. A porin antigen useful in these methods can be, for example, a protein immunoreactive with NANUC-2 and having at least 65% amino acid identity with SEQ ID NO: 28, SEQ ID NO: 29 or SEQ ID NO: 30.
In addition, there is provided a porin antigen, or tolerogenic fragment thereof, combined with a tolerogizing molecule. In a composition of the invention, a porin antigen can be, for example, a protein immunoreactive with NANUC-2 and having at least 65% amino acid identity with SEQ ID NO: 28, SEQ ID NO: 29 or SEQ ID NO: 30.
The present invention further provides methods of diagnosing UC by obtaining a sample from a subject suspected of having inflammatory bowel disease; contacting the sample with a Bacteroides antigen, or pANCA-reactive fragment thereof, under conditions suitable to form a complex of Bacteroides antigen, or pANCA-reactive fragment thereof, and antibody to Bacteroides antigen; and detecting the presence or absence of the complex, where the presence of the complex indicates that the subject has ulcerative colitis. A Bacteroides antigen useful in these methods can be, for example, a Bacteroides cecae protein immunoreactive with NANUC-2 and having a molecular weight of about 100 kDa by SDS-PAGE electrophoresis.
The invention also provides methods of inducing tolerance in a pANCA-positive patient with UC by administering to the patient an effective dose of a Bacteroides antigen, or tolerogenic fragment thereof. Such a Bacteroides antigen useful in these methods can be, for example, A Bacteroides cecae protein immunoreactive with NANUC-2 and having a molecular weight of about 100 kDa by SDS-PAGE electrophoresis.
Additionally, the invention provides a composition containing a Bacteroides antigen, or a tolerogenic fragment thereof, combined with a tolerogizing molecule. A Bacteroides cecae protein immunoreactive with NANUC-2 and having a molecular weight of about 100 kDa by SDS-PAGE electrophoresis is an example of a Bacteroides antigen useful in a composition of the invention.
Further provided herein are methods for identifying an agent useful for treating UC. The methods involve obtaining a sample of enteric bacteria from a patient with UC; isolating from the sample a bacterial species that expresses a pANCA-reactive antigen; contacting the bacterial species with an agent; and assaying for the reduced growth or viability of the bacterial species, where the reduced growth or viability of the bacterial species indicates that the agent is an agent useful for treating UC. Enteric bacterial species useful in these methods can be, for example, from the genera of Mycobacteria, Escherichia or Bacteroides.